Harvard College

University of California

Agilent Technologies, Inc.

Other

Government of the United States of America, As Represented by the Secretary of Commerce

Daniel Branton

Jene A Golovchenko

David W Deamer

Mark Albert Akeson

Gavin M King

Timothy J Denison

Jeffrey Robert Sampson

Amit Meller

Eric Brandin

Slaven Garaj

Gregor M Schurmann

George M Church

Jiali Li

Stephen Jordan Fleming

Michael J Aziz

John J Kasianowicz

Christopher John Russo

Ciaran J McMullan

Alexis Sauer

Warren J MoberlyChan

Alexis Sauer-Budge

Damon B Farmer

Roy Gerald Gordon

William Dunbar

Noah A Wilson

Haibing Peng

Roger Jinteh Arrigo Chen

Derek M Stein

Richard Baldarelli

Marc H Gershow

Seico Benner

Gregor M Schürmann

Kate Lieberman

Toshiyuki Mitsui

Seiko L Endo

Robin Abu-Shumays

Nicholas Hurt

Anpan Han

Dimitar M Vlassarev

El-Hadi S Sadki

Peng Chen

William B Dunbar

Garaj Slaven

Ciaran J Mcmullan

In a method for controlling translocation of a target polymer molecule through a nanopore, a clamp is reversibly bound at a site along the target polymer molecule length; the clamp and the target polymer molecule are disposed in an ionic solution that is in fluidic communication with a nanopore having an aperture diameter less than an outer diameter of the clamp. A constant translocation force is applied across the nanopore to induce travel of the target polymer molecule into the nanopore such that the reversibly bound clamp abuts the nanopore. A voltage pulse is applied across the nanopore that advances the target polymer molecule into the nanopore by one nucleotide, without either of chemical fuel and biochemical fuel provided to the clamp. The voltage pulse is repeatedly applied to cause a plurality of nucleotides to translocate through the nanopore. An indication of each nucleotide can be acquired during nucleotide translocation.

Deterministic control of polymer molecule translocation through a nanopore

A nanopore system provided herein includes first and second fluidic reservoirs in fluidic communication with a nanopore forming a fluidic path between the reservoirs. An enzyme clamp, provided in the first fluidic reservoir, abuts the nanopore and is reversibly bound to a sequential plurality of polymer subunits of a target polymer molecule in ionic solution. The clamp has an outer clamp diameter that is greater than the nanopore diameter. An electrical circuit includes an electrode in each of the reservoirs for applying a voltage bias across the nanopore. A pulse generator is connected in the electrical circuit to apply control pulses across the nanopore to step the clamp along sequential polymer subunits of the target polymer molecule. The system includes no fuel or source of fuel for the clamp. A controller is connected in the electrical circuit for controlling the collection of electrical indications of polymer subunits.

Deterministic stepping of polymers through a nanopore

In a method p for controlling translocation of a target polymer molecule through a nanopore, a clamp is reversibly bound to a sequential plurality of polymer subunits along the target polymer molecule length and the molecule and clamp are disposed in an ionic solution that is in fluidic communication with the nanopore. A constant translocation force is applied across the nanopore to induce travel of the target polymer molecule into the nanopore, until the clamp abuts the nanopore aperture and stops further travel of the target polymer molecule into the nanopore. Then a voltage control pulse is applied across the nanopore and/or a thermal control pulse is applied at the nanopore, with a pulse duration that steps the clamp along the target polymer molecule by no more than one polymer subunit in a direction opposite that of travel into the nanopore. No fuel is provided to the clamp.

There is provided a nanometric structure that includes a self-supporting nanometric material having a thickness of no more than about 5 nm. A plurality of nanopores is provided in the nanometric material, and the nanopore plurality has a density of at least about 1000 nanopores/cm. Each nanopore in the plurality of nanopores has a diameter that is no greater than about 10 nm. The plurality of nanopores is monodisperse in diameter with a variation of about &#xb1;30%. In a further nanometric structure provided herein there is included a self-supporting nanometric material having a thickness of no more than about 5 nm. A plurality of nanopores in the nanometric material includes at least about 50 nanopores. Each nanopore in the plurality of nanopores has a diameter that is no greater than about 10 nm. The plurality of nanopores is monodisperse in diameter with a variation of about &#xb1;30%.

Controlled fabrication of nanopores in nanometric solid state materials

There is provided a substantially bare, self-supported single-layer graphene membrane including a nanopore extending through a thickness of the graphene membrane from a first to a second membrane surface opposite the first graphene membrane surface. A connection from the first graphene membrane surface to a first reservoir provides, at the first graphene membrane surface, a species in an ionic solution to the nanopore, and a connection from the second graphene membrane surface to a second reservoir is provided to collect the species and ionic solution after translocation of the species and ionic solution through the nanopore from the first graphene membrane surface to the second graphene membrane surface. An electrical circuit is connected on opposite sides of the nanopore to measure flow of ionic current through the nanopore in the graphene membrane.

Nanometric material having a nanopore enabling high-sensitivity molecular detection and analysis

Provided is a solid state support structure including an aperture having a molecular entrance and a molecular exit. A first reservoir is in fluidic communication with the molecular entrance of the aperture and contains a molecule-bearing liquid solution. A second reservoir is in fluidic communication with the molecular exit of the aperture for containing a molecule-bearing liquid solution. A first liquid channel is connected to the first reservoir within less than about 300 microns of the aperture in the support structure and includes molecule-bearing liquid solution for delivery to the first reservoir. A second liquid channel is connected to the second reservoir for accepting molecule-bearing liquid solution from the second reservoir. An electrical connection between the first reservoir and the second reservoir imposes an electrical bias between the first reservoir and the second reservoir for driving the molecule-bearing liquid solution through the aperture in the solid state support structure.

Molecular characterization device

Devices and methods that can detect and control an individual polymer in a mixture is acted upon by another compound, for example, an enzyme, in a nanopore are provided. The devices and methods also determine (&#x2dc;&#x3e;50 Hz) the nucleotide base sequence of a polynucleotide under feedback control or using signals generated by the interactions between the polynucleotide and the nanopore. The invention is of particular use in the fields of molecular biology, structural biology, cell biology, molecular switches, molecular circuits, and molecular computational devices, and the manufacture thereof.

Compositions, devices, systems, and methods for using a nanopore

The invention features the use of graphene, a one atom thick planar sheet of bonded carbon atoms, in the formation of artificial lipid membranes. The invention also features the use of these membranes to detect the properties of polymers (e.g., the sequence of a nucleic acid) and identify transmembrane protein-interacting compounds.

Graphene supported artificial membranes and uses thereof

In a method of forming a nanopore in a nanometric material, a nanopore nucleation site is formed at a location that is interior to lateral edges of the nanometric material by directing a first energetic beam, selected from the group of ion beam and neutral atom beam, at the interior location for a first time duration that imposes a first beam dose which causes removal of no more than five interior atoms from the interior location to produce at the interior location a nanopore nucleation site having a plurality of edge atoms. A nanopore is then formed at the nanopore nucleation site by directing a second energetic beam, selected from the group consisting of electron beam, ion beam, and neutral atom beam, at the nanopore nucleation site with a beam energy that removes edge atoms at the nanopore nucleation site but does not remove bulk atoms from the nanometric material.

Provided is a first reservoir for containing a liquid solution including a molecule to be characterized and a second reservoir for containing a liquid solution. A solid state support includes a nanopore having a molecular inlet providing a fluidic connection to the first reservoir and a molecular outlet providing a fluidic connection to the second reservoir. An electrical connection is disposed between the first and second reservoirs to apply a molecular translocation voltage across the nanopore between the molecular inlet entrance and outlet exit. At least one electrical probe is disposed at the nanopore to apply a first voltage bias with respect to translocation voltage to slow progression of a molecule through the nanopore between the molecular inlet and outlet and to apply a second voltage bias with respect to translocation voltage to cause the molecule to proceed through the nanopore between the molecular inlet and outlet.

Molecular characterization with molecular speed control

The invention relates to a method for characterizing a target polynucleic acid by providing a surface containing a channel of a dimension sufficient to allow sequential monomer-by-monomer passage of a single-stranded polynucleic acid, but not of a double-stranded polynucleic acid; providing a source of hybridized target polynucleic acid at the surface; inducing passage of the target polynucleic acid through the channel, whereby the target polynucleic acid undergoes base pair separation (melts) prior to its passage; and making one or more measurements over time as the target polynucleic acid moves relative to the channel yielding data suitable to determine a monomer-dependent characteristic of the target polynucleic acid.

Characterization of individual polymer molecules based on monomer-interface interactions

Nanopore device with graphene supported artificial lipid membrane

In a method for imaging a solid state substrate, a vapor is condensed to an amorphous solid water condensate layer on a surface of a solid state substrate. Then an image of at least a portion of the substrate surface is produced by scanning an electron beam along the substrate surface through the water condensate layer. The water condensate layer integrity is maintained during electron beam scanning to prevent electron-beam contamination from reaching the substrate during electron beam scanning. Then one or more regions of the layer can be locally removed by directing an electron beam at the regions. A material layer can be deposited on top of the water condensate layer and any substrate surface exposed at the one or more regions, and the water condensate layer and regions of the material layer on top of the layer can be removed, leaving a patterned material layer on the substrate.

Electron beam processing with condensed ice

Systems and methods for analysis of polymers, e.g., polynucleotides, are provided. The systems are capable of analyzing a polymer at a specified rate. One such analysis system includes a structure having a nanopore aperture and a molecular motor, e.g., a polymerase, adjacent the nanopore aperture.

Methods and apparatus for characterizing polynucleotides

Growing community of inventors

Lexington, MA, United States of America

Daniel Branton